The objectives of the proposed research are: a) To develop methods to determine the solution properties of amino acid residues at active sites in both free enzymes and enzyme-substrate complexes. b) To identify the role of structural groups other than the aldehyde and amino groups in the coenzyme in enzyme-coenzyme relationships and even their plausible implication in enzymatic catalysis. c) To characterize those amino acids constituting the active center of selected examples of phosphopyridoxal dependent enzymes. d) To clarify the role of those amino acids (point c) in enzyme-coenzyme-substrate relationships and the mechanism of catalysis. e) To gain insight into possible changes in tertiary or quaternary protein structure during catalysis. f) To detect the effects elicited by interacting proteins on the microenvironment of strategic protein regions of the PLP-dependent protein or on groups within the bound pyridoxal phosphate (PLP) or pyridoxamine phosphate (PMP) molecules. g) To bring about new methodology for the selective use of PLP as a specific modifying agent of other proteins in solution or membranes. h) To exploit the preference for PLP by non PLP-requiring proteins to further our knowledge of localized perturbations resulting from ligand interactions with the protein during the heterotopic protein-protein interactions.